Nucleic acid chemical modifications
View Patent ↗The present invention provides nucleosides of formula (1) and oligonucleotides comprising at least one nucleoside of formula (2): Another aspect of the invention relates to a method of inhibiting the expression of a gene in cell, the method comprising (a) contacting an oligonucleotide of the invention with the cell; and (b) maintaining the cell from step (a) for a time sufficient to obtain degradation of the mRNA of the target gene.
1. A nucleoside represented by formula (3):
or isomers thereof, wherein:
one of D 1 and D 2 is H or a hydroxyl protecting group and the other of D 1 and D 2 is H, a hydroxyl protecting group, or a reactive phosphorus group; with the proviso that when one of D 1 and D 2 is H, the other of D 1 and D 2 is either H or a hydroxyl protecting group;
B is selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, a natural nucleobase, and a universal base;
Q is selected from the group consisting of O, S, NR 10 , and CH 2 ;
W is absent, O, S or NR′;
R 100 and R 200 are each independently selected from the group consisting of hydrogen, unsubstituted and substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
Z is OR 10 ;
R 20 and R 30 are each independently selected from the group consisting of hydrogen, acyl, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, 5-14 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-14 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, or 5-14 membered heterocyclic,
wherein, in each occurrence, the substituent group on the substituted C 1 -C 24 aliphatic is selected from the group consisting of halo, hydroxy, oxo, nitro, haloalkyl, alkyl, alkaryl, aryl, aralkyl, alkoxy, aryloxy, amino, acylamino, alkylcarbamoyl, arylcarbamoyl, aminoalkyl, alkoxycarbonyl, carboxy, hydroxyalkyl, alkanesulfonyl, arenesulfonyl, alkanesulfonamido, arenesulfonamido, aralkylsulfonamido, alkylcarbonyl, acyloxy, cyano, and ureidos,
provided that at least one R 100 , R 200 , R 300 , and R 400 is methyl.
2. The nucleoside of claim 1 , represented by formula (5):
or isomers thereof, wherein:
one of D 1 and D 2 is H, or a hydroxyl protecting group and the other of D 1 and D 2 is H, a hydroxyl protecting group, or a reactive phosphorus group; with the proviso that when one of D 1 and D 2 is H, the other of D 1 and D 2 is either H or a hydroxyl protecting group;
B is selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, a natural nucleobase, and a universal base;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
Z is OR 10 ;
R 20 and R 30 for each occurrence are independently selected from the group consisting of hydrogen, acyl, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, 5-14 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-14 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, an unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, or 5-14 membered heterocyclic.
3. The nucleoside of claim 1 , wherein:
B is a natural nucleobase or a universal base;
Q is O;
W is O;
R 100 and R 200 are each independently selected from the group consisting of hydrogen, C 1 -C 10 aliphatic, aryl, halogen, 5-8 membered heteroaryl, and 5-8 membered heterocyclic;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, C 1 -C 10 aliphatic, aryl, halogen, 5-8 membered heteroaryl, and 5-8 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
R 20 and R 30 for each occurrence are independently selected from the group consisting of hydrogen, acyl, C 1 -C 10 aliphatic, aryl, 5-8 membered heteroaryl, 5-8 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-8 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, an C 1 -C 10 aliphatic, aryl, 5-8 membered heteroaryl, or 5-8 membered heterocyclic.
4. The nucleoside of claim 3 , wherein:
R 100 and R 200 are each independently selected from the group consisting of hydrogen, C 1 -C 10 alkyl, aryl, and 5-8 membered heteroaryl;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, C 1 -C 10 alkyl, aryl, and 5-8 membered heteroaryl; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl; and
R 10 and R 10 ′ are each independently hydrogen or C 1 -C 10 alkyl.
5. The nucleoside of claim 1 , wherein the nucleoside has the formula of
wherein B is U, T, C Ac , A Bz , or G ibu .
6. An oligonucleotide comprising at least one nucleoside of formula (4):
or isomers thereof, wherein:
B is selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, a natural nucleobase, and a universal base;
Q is selected from the group consisting of O, S, NR 10 , and CH 2 ;
W is absent, O, S or NR′;
R 100 and R 200 are each independently selected from the group consisting of hydrogen, unsubstituted and substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
Z is OR 10 ;
R 20 and R 30 are independently selected from the group consisting of hydrogen, acyl, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, 5-14 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-14 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, or 5-14 membered heterocyclic,
wherein, in each occurrence, the substituent group on the substituted C 1 -C 24 aliphatic is selected from the group consisting of halo, hydroxy, oxo, nitro, haloalkyl, alkyl, alkaryl, aryl, aralkyl, alkoxy, aryloxy, amino, acylamino, alkylcarbamoyl, arylcarbamoyl, aminoalkyl, alkoxycarbonyl, carboxy, hydroxyalkyl, alkanesulfonyl, arenesulfonyl, alkanesulfonamido, arenesulfonamido, aralkylsulfonamido, alkylcarbonyl, acyloxy, cyano, and ureidos,
provided that at least one of R 100 , R 200 , R 300 and R 400 is methyl.
7. The oligonucleotide of claim 6 , wherein the nucleoside of formula (4) is represented by a nucleoside of formula (6):
or isomers thereof, wherein:
B is selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, a natural nucleobase, and a universal base;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, halogen, 5-14 membered heteroaryl, and 5-14 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
Z is OR 10 ;
R 20 and R 30 for each occurrence are independently selected from the group consisting of hydrogen, acyl, unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, 5-14 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-14 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, an unsubstituted or substituted C 1 -C 24 aliphatic, aryl, 5-14 membered heteroaryl, or 5-14 membered heterocyclic.
8. The oligonucleotide of 6 , wherein the nucleoside of formula (2) is selected from the group consisting of:
and isomers thereof, wherein:
R 1 , R 2 , and R 3 are each independently selected form the group consisting of C 1 -C 10 alkyl, aryl, C 3 -C 12 cycloalkyl, 5-14 membered heteroaryl, heterocycloalkyl and acyl;
B is an optionally substituted natural or non-natural nucleobase or a universal nucleobase;
X is O, S, or BH 3 ;
Y is O; and
m, n and p are each independently an integer from 1 to 10.
9. The oligonucleotide of 6 , wherein said oligonucleotide further comprises at least one non-phosphodiester backbone linkage.
10. The oligonucleotide of claim 9 , wherein the non-phosphodiester of said non-phosphodiester backbone linkage is selected from the group consisting of phosphorothioate, phosphorodithioate, alkyl-phosphonate and phosphoramidate.
11. The oligonucleotide of claim 9 , the non-phosphodiester backbone linkage is placed (a) between nucleosides of positions 1 and 2, and/or (b) between the 3′-terminal nucleosides.
12. The oligonucleotide of 6 , wherein said oligonucleotide further comprises at least one ligand bonded to a nucleobase, a sugar moiety, an internucleosidic linkage, or a 3′ or 5′ end of the oligonucleotide.
13. The oligonucleotide of 6 , wherein said oligonucleotide is a double-stranded oligonucleotide comprising a first strand and a second strand.
14. The oligonucleotide of claim 13 , wherein the nucleoside of formula (4) is present in either the first strand or the second strand.
15. The oligonucleotide of claim 14 , wherein said first strand is the sense strand.
16. The oligonucleotide of claim 14 , wherein said second strand is the antisense strand.
17. The oligonucleotide of claim 14 , wherein the nucleoside of formula (4) is present in both strands.
18. The oligonucleotide of 6 , wherein said oligonucleotide has a hairpin structure.
19. The oligonucleotide of 6 , wherein said oligonucleotide is an antisense, an antagomir, a microRNA, a siRNA, a pre-microRNA, an antimir, a ribozyme, RNA activator, U1 adaptor, immune stimulator or an aptamer.
20. The oligonucleotide of claim 19 , wherein said oligonucleotide is a single-stranded siRNA.
21. The oligonucleotide of claim 6 , wherein:
B is a natural nucleobase or a universal base;
Q is O;
W is O;
R 100 and R 200 are each independently selected from the group consisting of hydrogen, C 1 -C 10 aliphatic, aryl, halogen, 5-8 membered heteroaryl, and 5-8 membered heterocyclic;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, C 1 -C 10 aliphatic, aryl, halogen, 5-8 membered heteroaryl, and 5-8 membered heterocyclic; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl, thiocarbonyl, ═NR 20 , ═N—NR 20 R 30 , or ═N—NR 20 C(O)R 10 ;
R 20 and R 30 for each occurrence are independently selected from the group consisting of hydrogen, acyl, C 1 -C 10 aliphatic, aryl, 5-8 membered heteroaryl, 5-8 membered heterocyclic, OR 10 , COR 10 , CO 2 R 10 , and NR 10 R 10 ′; or R 20 and R 30 are taken together with the atom to which they are bound to form a 5-8 membered heterocyclic ring; and
R 10 and R 10 ′ are each independently hydrogen, C 1 -C 10 aliphatic, aryl, 5-8 membered heteroaryl, or 5-8 membered heterocyclic.
22. The oligonucleotide of claim 21 , wherein:
R 100 and R 200 are each independently selected from the group consisting of hydrogen, C 1 -C 10 alkyl, aryl, and 5-8 membered heteroaryl;
R 300 and R 400 are each independently selected from the group consisting of hydrogen, C 1 -C 10 alkyl, aryl, and 5-8 membered heteroaryl; or R 300 and R 400 are taken together with the atom to which they are bound to form a carbonyl; and
R 10 and R 10 ′ are each independently hydrogen or C 1 -C 10 alkyl.
23. The oligonucleotide of claim 6 , wherein the nucleoside has the formula of
wherein B is U, T, C Ac , A Bz , or G ibu .